Flagellated bacteria are common pathogens of humans and cause many important diseases including typhoid and cholera. Flagellins are dominant target antigens of the immune response during many intestinal bacterial infections and also in patients suffering from inflammatory bowel disease. Understanding the extraordinary ability of flagellins to activate innate and adaptive immune responses is therefore of critical importance for translational research on vaccines and adjuvants for intestinal infections and for basic research on the prevention of inflammatory disease. Using 2-year ARRA funding, we have found that the innate receptor TLR5 is essential for the development of a robust T cell response to flagellin, but that this requires an unconventional signaling pathway that has not previously been examined. In addition, we have now developed sensitive in vitro and in vivo assays to monitor the activation of flagellin-specific T cell responses by dendritic cells. In this renewal application we propose to build upon our successful studies completed with ARRA funding and continue studying the role of TLR5 in the development of flagellin-specific immunity. The specific aims of this sub-project are: Aim 1. Examine a new signaling pathway downstream of TLR5 to test the hypothesis that components of CLR and autophagic pathways are required by DCs to activate flagellin-specific T cells. Aim 2. Specific Aim 2. Examine the ability of intestinal DC subsets to activate flagellin-specific T cells, to test the hypothesis tha CCR6+ PP DCs are responsible for adaptive immune responses to flagellated pathogens. Our preliminary data describe the development of state-of-the-art technology to examine flagellin- specific T cell responses in vitro and in vivo. Our overall hypothesis is that TLR5 functions as a scavenger receptor on intestinal DCs to focus T cell responses onto flagellin. PUBLIC HEALTH RELEVANCE: Flagellins are bacterial proteins that become dominant target antigens in many intestinal infections and in patients suffering from inflammatory bowel disease. Our proposal aims to examine how bacterial flagellins interact with the innate immune system and in particular understand how the innate and adaptive immune response to flagellin is functionally integrated. Understanding why flagellins are particularly immunogenic is an important issue for the development of new vaccines against flagellated microbes, the current use of flagellins as vaccine adjuvants, and for the potential to develop new therapies targeting intestinal inflammation against enteric bacteria.